protegrin

Certainly, the striking correlation with the extent of protegrin binding must be one of these.

Why did fewer protegrin molecules bind to B. cepacia than to P. aeruginosa ?

We also found that at any given peptide concentration, many fewer protegrin molecules bound to intact B. cepacia and its lipid A than to the corresponding P. aeruginosa preparations.

Moreover, in addition to binding the LPS or lipid A molecules we added to the well, some of the initially added protegrin could diffuse radially into the underlay gel.

If most of the P. aeruginosa lipid A phosphates are unmodified, and if these phosphates are principal protegrin binding sites, then one would expect exactly the results shown in Fig.

Thus, it is a reasonable inference that the relative resistance of B. cepacia to PG-1 is a consequence of the smaller number of protegrin molecules bound per unit area of bacterial surface.

In model systems, the ability of protegrins to permeabilize membranes shows pronounced "concentration-gating", an indication that it is influenced by the density of bound protegrin molecules per unit area of membrane [ 47 ] .

Although the acyl chains of lipid A might also provide alternative binding sites for protegrin, the orientation of lipid A on the HPA sensor chip ("butter-side down") probably removes them from consideration, at least under our study conditions.

This protegrin variant differed from PG-1 in two ways: Phe7 replaced Tyr7 in the protegrin domain, and an N-terminal glycine-rich hexapeptide extension (GGGYGG) with a single tyrosine residue was present.

Although up to one third more protegrin molecules bound to P. aeruginosa LPS than to B. cepacia LPS at protegrin concentrations of 1-6 μM, these differences disappeared when we tested higher PG-1 concentrations.

By mixing a constant amount of protegrin (within this linear range) with graded amounts of purified LPS and lipid A, we could readily determine how much of these ligands were needed to reduce the activity of 50 ng of PG-1 by 50% (the EC 50 values).

The acyl chains of lipid A form an integral part of the outer membrane bilayer, and electrostatic or hydrophobic binding of protegrin to lipid A could easily disturb the organization and acyl-chain packing of the outer membrane, both vital in maintaining its integrity and barrier function.

To resolve these seemingly discrepant results, it is important to recall that in SPR assays, the sensor chips were constantly bathed with a fixed protegrin concentration ranging from 1-20 μM and that in radial diffusion assays, the wells received a fixed initial amount of protegrin (50 ng).